Journal of Natural Products Vol. 58, No. 1 , p p . 134-137,January 1995
134
OBOVATINE, A NEW BISINDOLE ALKALOID FROM STEMMADENIA OBOVATA E. VALENCIA,A. MADINAVEITM, J. BERMEJO,*and A.G. GONZ~LEZ Centro a2 Productos Naturales Orgdnicos “Antonio Gonz6la,” IPNAC-CSIC, La Laguna, Tenerqe, Canary Islands, Spain ABSTRACT.-A new bisindole alkaloid, obovatine El],has been isolated from the leaves of Stmmadenia obovata together with theknowncompounds, bis[l l-hydroxycoronaridin-12-y1][23, 1 1-hydroxycoronaridine [51, and voacristine 161.The structure of 1 was established by spectroscopic analysis and chemical transformations.
Stemmadenzaobovata (Benth.) Woods. (Apocynaceae),is one of ten species (1-7) known to produce ibogaine-type indole alkaloids (8). This particular species has no place in the popular pharmacopeia, unlike, for instance,S. donnelsmitbii, which is widely used in Central America to treat rheumatism, eye inflammation, and toothache (9). Alkaloids were first obtained from the bark, fruit and seeds ofS. obovuta by Collera et ul. in 1962 (2) and now a new bisindole alkaloid, obovatine 117, has been isolated from the leaves of
this species, together with the known alkaloids, bisll l-hydroxycoronaridin-12yl} [2} (10), 11-hydroxycoronaridine [ 5 } (1l), and voacristine (19-hydroxyvoacangine) 161 (12). The CHC1, extract of the leaves of S. obovata was subjected to a combination of SephadexLH-20 and Si gel chromatography to afford four ibogaine-type indole alkaloids, 1, 2, 5 , and 6 . Compound 1 proved to be a new indole alkaloid dimer, obovatine. Compound 1, C43H52N406, amor-
18
1C
1 R,=H,R,=Me 2
R,=R,=H
3 R,=R,=H 3 R,=R,=Ac 4 R,=R,=Me
COOCHJ
5 R,=R,=H, R,=OH
6 R,=OMe, R,=H, R,=OH
January 19951 Valencia et al. : Bisindole Alkaloids from Stmmadmia phous alkaloid, {ol}25D +17.8°(c=0.13, CHCl,), had a molecular ion peak at mlz 720 (100%) and significant fragment ions at mlz 360 (13%), 208 (5%), 136 (56%), and 135 (13%) characteristic of an ibogaine-type alkaloid (13). The uv spectrum showed absorptions for a substituted indole-type ring, and a bathochromic shift, after base addition, indicated the presence of a phenolic hydroxy group in the indole ring. Ir bands appeared at 3540 cm-' for OH, 3340 -1 cm for NH, and at 1720 cm-' for a carbonyl ester. The structure of 1 was determined by comparison of its 'H- and l3C-nmr data with those of other ibogaine alkaloids, notably 2, a symmetrical alkaloid isolated from Bonafousia tetrastachya ( H u m b o l d t , Bonpland et K u n t h ) Markgraf (Apocynaceae) (10). The 400 MHz 'H-nmr spectrum of 1 revealed the presence of two sets ofwell-defined orthocoupled aromatic protons at 6 7.52 and 6.91 (J=8.5 Hz) and 6 7.42 and 6.98
(J=8.5 Hz), in consonance with an indole nucleus. The exact chemical shifts of the aromatic moiety and the C- 12/C-12 ' linkage were ascertained by a long-range heteronuclear multiple bond correlation spectroscopy (HMBC) 2D nmr experiment (Table 1). The presence of two three-proton singlets at 6 3.67 and 3.64 assigned to carbomethoxy groups indicated a dimeric molecule, as did the two N H group singlets at 6 7.37 and 7.35. Proton-carbon chemical shift correlations for all the carbons directly bonded to protons could be established in an HMQC experiment (Table 2).
H
TABLE 2. Nmr Data of 1 16 (pi
2,2". . . . . . . . . . . 3,3'". . . . . . . . . . . 5,5". . . . . . . . . . . 6,6". . . . . . . . . . . 7,7" . . . . . . . . . .
-
83'
Correlated C
1).
CDCIJ.
DEPT
Position
...........
9,9' . . . . . . . . . . . 10,lO' . . . . . . . . . 11,ll' . . . . . . . . . 12,12' . . . . . . . . . 13,13' . . . . . . . . . 14,14". . . . . . . . . 15,15". . . . . . . . . 16,16". . . . . . . . . 17,17". . . . . . . . . 18,18". . . . . . . . . 19,19". . . . . . . . . 20,20". . . . . . . . . 21,21". . . . . . . . .
c=o . . . . . . . . . .
OCH, OCH, (Ar) . . . . . . NH . . . . . . . . . . .
137.0, 135.4 52.6, 52.4 5 3.1 (double) 22.1 (double) 110.7 (double) 124.4, 123.1 110.1, 119.4 106.2, 119.3 149.4, 153.2 103.1, 102.7 134.2, 134.4 27.4, 27.3 32.0, 31.9 55.1, 55.0 36.5, 36.1 11.7, 11.6 26.8, 26.7 39.0, 39.0 58.2, 57.2 175.4, 175.0 52.3, 52.0 57.0
135
C
CH2 CH, CH, C C CH CH C C C CH CH2 C CH2 Me CH, CH CH C Me Me
2.90 (4H, sharp dd) 3.40, 3.23 (2H each, m) 3.07, 3.27 (2H each, m)
7.42, 7.52 (1H each, d,J=8.5 Hz) 6.98, 6.91 (1H each, d,J=8.5 Hz)
1.87 (2H, m) 1.71, 1.13 (2H each, m) 1.87, 2.44 (2H each, m) 0.89 (6H, dt,]=7.1 Hz) 1.56, 1.43 (2H each, m) 1.29 (2H, m) 3.54 (2H, m) 3.67, 3.64 (3H each, s) 3.79 (3H, s) 7.37, 7.35 (1H each, s)
r h e assignments for designated pairs o arbon and proton signals can be interchanged.
Journal of Natural Products
136
The 13C-nmr spectrum (100 MHz) exhibited 40 signals for the 4 3 carbon atoms of the molecule, with three corresponding to more than one carbon. The DEPT spectrum revealed the presence of two sp3quaternary carbons, fourteen sp2 quaternary carbons (one of which represented two carbons), six sp3methine carbons (two of two carbons apiece), four sp2 carbon hydrogens, twelve sp3 methylenes, and five methyl groups (Table 2). The signals for two sp2 quaternary carbons at 6 149.4 and 153.2 indicated hydroxy and methyl groups attached to their respective aromatic rings, in good agreement with the four sp2CH signals. The appearance of four low-field sp3methylenecarbonsat652.5,52.4,and53.1 (double) and two methine carbons at 6 57.2 and 58.1 suggested links with a nitrogen atom. When compound 2 was methylated with CH2N2in Et,O, the monomethyl derivative 1 and the dimethyl derivative 4 were obtained, confirming chemically the structure of 1.Compound 4 has not been previously found in nature. EXPERIMENTAL GENERALEWERIMENTALPROCED~-Nmr
spectra were recorded in CDCI, for ‘H nmr at 400 and 200 MHz, and for ”C nmr at 100 and 50.32 MHz with TMS as internal standard. High- and low-resolution ms were run on a VG Micromass ZAB-2F spectrometer at 70 eV. Optical rotations were measured on a Perkin-Elmer model 241 polarimeterusingaNalampat25”. Irspectrawere obtained on a Nicolet 5PC Ft-ir spectrometer using CHCI,.Theuvspectrawere run on a HewlettPackard HP-8254-A diode array uv spectrophotometer. The gel filtration column (Sephadex LH20) used hexane-MeOH-CHCI, (2:l:l) as solvent while the eluents for tlc and Si gel cc were n-hexane or EtOAc or mixtures thereof. Dragendorff or Ehrlich reagents were used as tlc spray reagents. MATERLU.-haves of Stmmadeniu (Benth.) Woods. were collected by Mn. CarmenGaldamosinFebruary1993, intheareaof PlayadeMonagre, ProvinciadeLosSantos,Panama, and identified by Prof. Mireya Correa, Director of the Herbarium of the University of Panama. A voucher specimen (FLORPAN 1283) is filed with the Herbarium of the University of Panama. ohutu
EXTRACTION AND IWLATION.-Dried,
pow-
mol. 58, No. 1
dered leaves (678 g) were extracted with hot 90% EtOH. The solvent was evaporated in v u a o to a residue that was diluted in 5 % HCI to precipitate a deposit. After filtration of the mixture the solution was extracted with CHCI,. T h e aqueous layer was basified with “,OH to pH 10 and extracted exhaustively with CHCI, followed by drying (Na,SO,). Evaporation yielded a dark residue (2.9 g ) that was then chromatographed on a Sephadex LH-20 column with n-hexane-MeOH-CHCI, (2:l:l) as eluent, affording 1in pure form (10.2 mg); the ocher alkaloid fractions were combined and subjected to repeated cc on Si gel with nhexane and EtOAc mixtures of increasing polarity until only EtOAc was present. The alkaloid fractions were finally purified by prep. tlc to afford three amorphous compounds, 2,5, and 6 in 17.5, 6.5, and 7.8 mg yield, respectively. Obwutine Ill.-Amorphous, [(LID 17.8” ( ~ 0 . 1 3CHCI,); , hrms m/z found [MI’ 720.3886, C,,H,,N,O, requires 720.3886; uv (MeOH) A max(1og E) 234 (4.76), 304 (4.38) nm; fMeONa 238 (4.76), 306 (4.31) nm; ir (CHCI,) u max 3540,3340,1720cm-’; ms mlz 720 ([MI’, loo), 360 (13), 208 (5), 136 (56), 135 (13); ’H- and ”Cn m r data, see Table 2. Acetykution of2.-Ac20 (1 ml) was added to a solution of 2 (5.2 mg) in pyridine (1 ml) and the mixture stirred at room temperature overnight. The excess pyridine was removed under high vacuum pressure to give a residue (6.8 mg). T h e residue was chromatographed on Si gel to give 3 (5.0 mg) as an amorphous compound, [(LID 31.8” ( ~ 0 . 0 6CHCI,); , ‘H nmr (CDCI,, 200 MHz) 6 0.87 (6H, t,J=7.3 Hz, H,-18 and H3-18’), 1.15 (2H, m, H,-15 and Hs-15’), 1.31 (2H, m, H-20 and H-20’), 1.48 (4H, dq,J=7.0 Hz, H2-19and H2-19’), 1.78 (2H, m, H,-15 and HR-15’),1.83 (4H, m, H-14 and H-14’, and H,-17 and H,17’), 1.84 (6H, s, 2 XCOMe), 2.48 (2H,dJ= 13.0 Hz, H,-17 and Hs-17’), 2.87 (4H, m, H,-3 and H,-3‘), 3.10 (4H, m, H,-6 and H2-6’), 3.23 (2H, m, H,-5 and H,-5’), 3.38 (2H, m, H,-5 and H,5’), 3.53(2H,narrowd,J=l.l Hz,H-21 andH21’), 3.66(6H,s, 2XCOOMe), 6.93 and 7.00( 1H each, d,J=8.5 Hz, H-10 and H-lo’), 7.40 (2H, br s, ZXNH), 7.45 and 7.55 (1H each, d,J=8.5 Hz, H-9, H-9’); I3C n m r (CDCI,, 50.32 MHz) 6 11.62 (C-18andC-18’), 20.52 (2XCOMe), 22.18 (C-6andC-6’), 26.72 (C-19 andC-19’), 27.31 (C14andC-14’), 31.91 (C-15 andC-l5‘), 36.22(C17 and C-17’), 38.81 (C-20 andC-20‘), 51.74(C3 and C-3’), 52.64 (2XCOOMe), 53.05 (C-5 and C-5’), 55.06 (C-16 and C-16’), 57.75 (C-21 and C-21’), 108.62 (C-12 and C-12’), 110.72 (C-7 and C-7’), 114.55 (C-10 and C-lo’), 118.95 (C9 and C-9‘), 126.9O(C-8 andC-8’), 134.36(C-13 andC-l3‘), 137.63 (C-2 andC-2’), 144.32 (C-11 and C - l l ’ ) , 169.83 (2XCOMe), 174.77 (2XCOOMe); hrms m/z [MI’ 790.39415, calcd
January 19951 Valencia et al. : Bisindole Alkaloids from Stmmadenia for C46H14N408[MI’ 790.34416; ms mlz 790
{MI’ (65), 395 (1l), 208 (lo), 136 (100), 135 (3 1). MethyIation of 2.-An ethereal solution of CH,N, was added to a solution of 2 (10.1 mg) in Me,CO (3 ml) until the yellow color persisted and was then kept in the dark overnight. Removal of the solvent left a residue (12.7 mg) that showed two close running zones under tlc. These two compounds were separated by prep. tlc on Si gel using n-hexane and n-hexane-EtOAc (7:3), to obtain 1(7.4 mg) and 4 (3.1 mg). Dimethylobovatine {4f.-Amorphous compound, [ C X ~ D23.6’ ( ~ 0 . 2 7CHCI,); , hrms mlz [MI’ 734.4060, calcd for C,H,,N,O, [MI’ 734.4043; ms mlz 734 [MI’ (42), 367 (58), 208 (19), 136 (loo), 135 (40); ‘H n m r (CDCI,, 200 MHz) 6 0.89 (6H, dt,J=7.1 Hz, H,-18 and H,18’), 1.14(2H, m, H,-15 andH,-15‘), 1.25 (2H, m,H-20andH-20’), 1.60(4H,m,H,-19andHZ19’), 1.79(2H,m, H,-15 andH,-15’), 1.84(2H, m, H,-17 and HR-17’), 2.45 (2H, m, H,-17 and H,-17’),2.90(4H,m,H2-3andH2-3’), 3.07(2H, m, H,-6 and H,-6’), 3.23 (2H, m, H,-5 and H,5 ’ ) , 3.27 (2H, m, H,-6 and H,-6’), 3.40 (2H, m, H,-5 and H,-5’), 3.52 (2H, m, H-21 and H-21’),
3.65(6H,s,2XCOOMe),3.81(6H,s,ZXArOMe), 6.91 and 6.96 (1H each, d,J=8.3 Hz, H-10 and H-lo’), 7.40 (2H, br s, ZXNH), 7.42 and 7.51 (1H each, d, J=8.3 Hz, H-9 and H-9’). ACKNOWZEDGMENTS This work has been partly subsidized by Grant No. PB91-0049(CICYT)andtheConsejeria de Educaci6n de Canarias (P28/08.03.90). We should like to thank Prof. Dr. Mahabir P. Gupta, Pharmacognosy Research Unit (FLORPAN), University of Panama, Panama, for providing us with the plant material. A.G.G. is indebted to the Colegio Libre de EmCritos, Madrid, Spain.
137
LITERATURE CITED F. Walls, 0. Collera, and A. Sandoval L., Tetrahedron, 2, 173 (1958). 2. 0.Collera, F. Walls, A. Sandoval,F. Garcia, J. Herran, and M.C. Perezamador,BoI. Inst. Quim.Univ.N a d Auton. Mex., 1 4 , 3(1962); Chem. Abstr., 59, 1957d (1963). 3. J.A. Saenz and M. Nassar, Rev. Biol. Trop. Univ. Costa Rica, 13, 207 (1965); Chem. Abstr., 65, 10963h (1966). 4. X.A. Dominguez, D. Butruille, and P. Pereyra, Rev. Latinoam. Quim., 7,98 (1976); Chem. Abstr., 8 5 , 1195432 (1976). 5. J.F. Ciccio, V.H. Castro, and F. Acuna, An. Quim., Ser. C, 78,13 5 ( 1982); Chem. A bstr., 9 6 , 2 1 4 3 6 4 ~(1982). 6. J. Stkkigt, K.H. Pawelka, A. Rother, and B. Deuss,Z. Naturfosch., 37C, 857 (1982). 7. M.P. Gupta, D. Alvarez, P.N. Solis, M. Gwel, and H. Achenbach,Planta Med.,57, 502 (1991). 8. W.I. Taylor, in: “The Alkaloids. Chemistry and Physiology,” Ed. by R.H.F. Manske, Academic Press, New York, 1968, Vol. 11, pp. 80-81. 9. J.F. Morton, “Atlas of Medicinal Plants of Middle America,” Charles C. Thomas Publisher, Springfield, IL, 1981, p. 681. 10. M. Damak, C. Poupat, and A. Ahond, Tetrahedron Lett., 3531 (1976). 11. F. Ladhar, M. Damak,A. Ahond,C. Poupat, P. Potier, and C. Moretti,J. Nat. Prod., 44, 459 (1981). 12. T.R. Govindachari,B.S.Joshi, A.K. s a k s e ~ , S.S. Sathe, and N. Viswanathan, Tetrahedron Lett., 3873 (1965). 13. W.I. Taylor, in: “The Alkaloids. Chemistry and Physiology,” Ed. by R.H.F. Manske, Academic Press, New York, 1965, Vol. 8, p. 221. Received I 9 July I994 1.
